Stem Cell Fate Research Articles

Metabolic regulation of adipose stem cell fate and function: from bench to clinic

Metabolic regulation of adipose stem cell fate and function: from bench to clinic

Small extracellular vesicles of organoid-derived human retinal stem cells remodel Müller cell fate via miRNA: A novel remedy for retinal degeneration

Remodeling retinal Müller glial fate, including gliosis inhibition and pro-reprogramming, represents a crucial avenue for treating degenerative retinal diseases. Stem cell transplantation exerts effects on modulating retinal Müller glial fate. However, the optimized stem cell products and the underlying therapeutic mechanisms need to be investigated. In the present study, we found that retinal progenitor cells from human embryonic stem cell-derived retinal organoids (hERO-RPCs) transferred extracellular vesicles (EVs) into Müller cells following subretinal transplantation into RCS rats. Small EVs from hERO-RPCs (hERO-RPC-sEVs) were collected and were found to delay photoreceptor degeneration and protect retinal function in RCS rats. hERO-RPC-sEVs were taken up by Müller cells both in vivo and in vitro, and inhibited gliosis while promoting early dedifferentiation of Müller cells. We further explored the miRNA profiles of hERO-RPC-sEVs, which suggested a functional signature associated with neuroprotection and development, as well as the regulation of stem cell and glial fate. Mechanistically, hERO-RPC-sEVs might regulate the fate of Müller cells by miRNA-mediated nuclear factor I transcription factors B (NFIB) downregulation. Collectively, our findings offer novel mechanistic insights into stem cell therapy and promote the development of EV-centered therapeutic strategies.

A high-throughput screen in mESCs to identify the cross-talk between signaling, endocytosis, and pluripotency.

Embryonic stem cell fate is regulated by various cellular processes. Recently, the process of endocytosis has been implicated in playing a role in the maintenance of self-renewal and pluripotency of mouse embryonic stem cells. A previous siRNA-based screen interrogated the function of core components of the endocytic machinery in maintaining the pluripotency of embryonic stem cells, revealing a crucial role for clathrin mediated endocytosis. A number of other proteins involved in key signaling pathways have also been shown to both regulate and be regulated by endocytosis. We collated a list of 1141 genes in connection to the term "endocytosis" from Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO), excluding those previously interrogated, and examined the effect of their knockdown on the pluripotency of mouse embryonic stem cells (mESCs) using levels of green fluorescent protein driven by the Oct4 promoter. We used high-throughput screening followed by an automatedMATrix LABoratory(MATLAB)-based analysis pipeline and assessed changes in GFP fluorescence as a readout for ESC pluripotency. Through this screen we identified a number of genes, many hitherto not associated with stem cell pluripotency, which upon knockdown either resulted in a significant increase or decrease of GFP fluorescence. We further present validation for some of these hits. We present a workflow aimed to identify genes involved in signaling pathways which can be regulated by endocytosis, and that affect the pluripotency of ESCs.

Targeting adipocyte ESRRA promotes osteogenesis and vascular formation in adipocyte-rich bone marrow

Excessive bone marrow adipocytes (BMAds) accumulation often occurs under diverse pathophysiological conditions associated with bone deterioration. Estrogen-related receptor α (ESRRA) is a key regulator responding to metabolic stress. Here, we show that adipocyte-specific ESRRA deficiency preserves osteogenesis and vascular formation in adipocyte-rich bone marrow upon estrogen deficiency or obesity. Mechanistically, adipocyte ESRRA interferes with E2/ESR1 signaling resulting in transcriptional repression of secreted phosphoprotein 1 (Spp1); yet positively modulates leptin expression by binding to its promoter. ESRRA abrogation results in enhanced SPP1 and decreased leptin secretion from both visceral adipocytes and BMAds, concertedly dictating bone marrow stromal stem cell fate commitment and restoring type H vessel formation, constituting a feed-forward loop for bone formation. Pharmacological inhibition of ESRRA protects obese mice against bone loss and high marrow adiposity. Thus, our findings highlight a therapeutic approach via targeting adipocyte ESRRA to preserve bone formation especially in detrimental adipocyte-rich bone milieu.

Microbial metabolite steers intestinal stem cell fate under stress

Recently in Cell Metabolism, Wei etal.1 unveiled a brain-to-gut pathway that conveys psychological stress to intestinal epithelial cells, leading to their dysfunction. This gut-brain axis involves a microbial metabolite, indole-3-acetate (IAA), as a niche signal that hampers mitochondrial respiration to skew intestinal stem cell (ISC) fate.

Size-Optimized Layered Double Hydroxide Nanoparticles Promote Neural Progenitor Cells Differentiation of Embryonic Stem Cells Through the Regulation of M6A Methylation.

The committed differentiation fate regulation has been a difficult problem in the fields of stem cell research, evidence showed that nanomaterials could promote the differentiation of stem cells into specific cell types. Layered double hydroxide (LDH) nanoparticles possess the regulation function of stem cell fate, while the underlying mechanism needs to be investigated. In this study, the process of embryonic stem cells (ESCs) differentiate to neural progenitor cells (NPCs) by magnesium aluminum LDH (MgAl-LDH) was investigated. MgAl-LDH with diameters of 30, 50, and 100 nm were synthesized and characterized, and their effects on the cytotoxicity and differentiation of NPCs were detected in vitro. Dot blot and MeRIP-qPCR were performed to detect the level of m6A RNA methylation in nanoparticles-treated cells. Our work displayed that LDH nanoparticles of three different sizes were biocompatible with NPCs, and the addition of MgAl-LDH could significantly promote the process of ESCs differentiate to NPCs. 100 nm LDH has a stronger effect on promoting NPCs differentiation compared to 30 nm and 50 nm LDH. In addition, dot blot results indicated that the enhanced NPCs differentiation by MgAl-LDH was closely related to m6A RNA methylation process, and the major modification enzyme in LDH controlled NPCs differentiation may be the m6A RNA methyltransferase METTL3. The upregulated METTL3 by LDH increased the m6A level of Sox1 mRNA, enhancing its stability. This work reveals that MgAl-LDH nanoparticles can regulate the differentiation of ESCs into NPCs by increasing m6A RNA methylation modification of Sox1.

Redox regulation of meristem quiescence: outside/in.

Quiescence is an essential property of meristematic cells, which restrains the cell cycle while retaining the capacity to divide. This crucial process not only facilitates life-long tissue homeostasis and regenerative capacity but also provides protection against adverse environmental conditions, enabling cells to conserve the proliferative potency while minimizing DNA damage. As a survival attribute, quiescence is inherently regulated by the products of aerobic life, in particular reactive oxygen species (ROS) and the redox (reduction/oxidation) mechanisms that plant have evolved to channel these into pervasive signals. Adaptive responses allow quiescent cells to compensate for reduced oxygen tension (hypoxia) in a reversible manner, while the regulated production of the superoxide anion (O2·-) facilitates cell division and the maintenance of stem cells. Here we discuss the role of ROS and redox reactions in the control of the quiescent state in plant meristems, and how this process is integrated with cellular energy and hormone biochemistry. We consider the pathways that sense and transmit redox signals with a focus on the central significance of redox regulation in the mitochondria and nucleus, which is a major regulator of quiescence in meristems. We discuss recent studies that suggest that ROS are a critical component of the feedback loops that control stem cell identity and fate, and suggest that the ROS/hypoxia interface is an important 'outside/in' positional cue for plant cells, particularly in meristems.

Crosstalk between DNA Damage Repair and Metabolic Regulation in Hematopoietic Stem Cells.

Self-renewal and differentiation are two characteristics of hematopoietic stem cells (HSCs). Under steady physiological conditions, most primitive HSCs remain quiescent in the bone marrow (BM). They respond to different stimuli to refresh the blood system. The transition from quiescence to activation is accompanied by major changes in metabolism, a fundamental cellular process in living organisms that produces or consumes energy. Cellular metabolism is now considered to be a key regulator of HSC maintenance. Interestingly, HSCs possess a distinct metabolic profile with a preference for glycolysis rather than oxidative phosphorylation (OXPHOS) for energy production. Byproducts from the cellular metabolism can also damage DNA. To counteract such insults, mammalian cells have evolved a complex and efficient DNA damage repair (DDR) system to eliminate various DNA lesions and guard genomic stability. Given the enormous regenerative potential coupled with the lifetime persistence of HSCs, tight control of HSC genome stability is essential. The intersection of DDR and the HSC metabolism has recently emerged as an area of intense research interest, unraveling the profound connections between genomic stability and cellular energetics. In this brief review, we delve into the interplay between DDR deficiency and the metabolic reprogramming of HSCs, shedding light on the dynamic relationship that governs the fate and functionality of these remarkable stem cells. Understanding the crosstalk between DDR and the cellular metabolism will open a new avenue of research designed to target these interacting pathways for improving HSC function and treating hematologic disorders.

Temporal regulation of the Mediator complex during muscle proliferation, differentiation, regeneration, aging, and disease.

Genesis of skeletal muscle relies on the differentiation and fusion of mono-nucleated muscle progenitor cells into the multi-nucleated muscle fiber syncytium. The temporally-controlled cellular and morphogenetic changes underlying this process are initiated by a series of highly coordinated transcription programs. At the core, the myogenic differentiation cascade is driven by muscle-specific transcription factors, i.e., the Myogenic Regulatory Factors (MRFs). Despite extensive knowledge on the function of individual MRFs, very little is known about how they are coordinated. Ultimately, highly specific coordination of these transcription programs is critical for their masterfully timed transitions, which in turn facilitates the intricate generation of skeletal muscle fibers from a naïve pool of progenitor cells. The Mediator complex links basal transcriptional machinery and transcription factors to regulate transcription and could be the integral component that coordinates transcription factor function during muscle differentiation, growth, and maturation. In this study, we systematically deciphered the changes in Mediator complex subunit expression in skeletal muscle development, regeneration, aging, and disease. We incorporated our in vitro and in vivo experimental results with analysis of publicly available RNA-seq and single nuclei RNA-seq datasets and uncovered the regulation of Mediator subunits in different physiological and temporal contexts. Our experimental results revealed that Mediator subunit expression during myogenesis is highly dynamic. We also discovered unique temporal patterns of Mediator expression in muscle stem cells after injury and during the early regeneration period, suggesting that Mediator subunits may have unique contributions to directing muscle stem cell fate. Although we observed few changes in Mediator subunit expression in aging muscles compared to younger muscles, we uncovered extensive heterogeneity of Mediator subunit expression in dystrophic muscle nuclei, characteristic of chronic muscle degeneration and regeneration cycles. Taken together, our study provides a glimpse of the complex regulation of Mediator subunit expression in the skeletal muscle cell lineage and serves as a springboard for mechanistic studies into the function of individual Mediator subunits in skeletal muscle.

Enhancing the Maturation of Human Pluripotent Stem Cell-Derived Cardiomyocytes with an n-Type Organic Semiconductor Coating.

Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) are a promising cell source for cardiac regenerative medicine and in vitro modeling. However, hPSC-CMs exhibit immature structural and functional properties compared with adult cardiomyocytes. Various electrical, mechanical, and biochemical cues have been applied to enhance hPSC-CM maturation but with limited success. In this work, we investigated the potential application of the semiconducting polymer poly{[N,N'-bis(2-octyldodecyl)-naphthalene-1,4,5,8-bis(dicarboximide)-2,6-diyl]-alt-5,5'-(2,2'-bithiophene)} (P(NDI2OD-T2)) as a light-sensitive material to stimulate hPSC-CMs optically. Our results indicated that P(NDI2OD-T2)-mediated photostimulation caused cell damage at irradiances applied long-term above 36 μW/mm2 and did not regulate cardiac monolayer beating (after maturation) at higher intensities applied in a transient fashion. However, we discovered that the cells grown on P(NDI2OD-T2)-coated substrates showed significantly enhanced expression of cardiomyocyte maturation markers in the absence of a light exposure stimulus. A combination of techniques, such as atomic force microscopy, scanning electron microscopy, and quartz crystal microbalance with dissipation monitoring, which we applied to investigate the interface of the cell with the n-type coating, revealed that P(NDI2OD-T2) impacted the nanostructure, adsorption, and viscoelasticity of the Matrigel coating used as a cell adhesion promoter matrix. This modified cellular microenvironment promoted the expression of cardiomyocyte maturation markers related to contraction, calcium handling, metabolism, and conduction. Overall, our findings demonstrate that conjugated polymers such as P(NDI2OD-T2) can be used as passive coatings to direct stem cell fate through interfacial engineering of cell growth substrates.

Two‐Way Regulation of MgFe‐LDH on ESC Cell Fate via Temporal Selective Activation of Different Cell Membrane Receptors

AbstractStem cell fate regulation by biomaterials has considerable therapeutic potential; however, the mechanisms whereby biomaterials instruct stem cells for accurate fate regulation still need to be elucidated. Layered double hydroxide (LDH) nanoparticles are widely used for tissue regeneration because of their excellent physicochemical properties and biocompatibility. This study reveals that MgFe‐LDH has bidirectional regulatory effects on the differentiation of mouse embryonic stem cells into neural progenitor cells, essentially suppressing cell differentiation on the 1st day and promoting cell differentiation on the 3rd day. Single‐cell transcriptome sequencing and simulated computation of structural analysis demonstrate that MgFe‐LDH interacts with the membrane receptors leukemia inhibitory factor receptor (LIFR, 15 nm above the cell membrane) and pached1 (PTCH1, 6 nm above the cell membrane) on the 1st and 3rd day of differentiation, respectively. LIFR has more contact with MgFe‐LDH than PTCH1 and relatively weaker electrostatic interaction energies (−11010.0 and −9829.3 kJ mol−1 for PTCH1 and LIFR, respectively). These factors determine receptor binding and activation priority, followed by cell fate regulation. The results indicate that MgFe‐LDH regulate the activity and fate of stem cells dually, thus creating possibilities for personalized therapy in neurodevelopment and regenerative medicine through an in‐depth study of cell membrane receptors.

Nuclear curvature determines Yes-associated protein localization and differentiation of mesenchymal stem cells

Controlling mesenchymal stem cell (MSC) differentiation remains a critical challenge in MSCs’ therapeutic application. Numerous biophysical and mechanical stimuli influence stem cell fate; however, their relative efficacy and specificity in mechanically directed differentiation remain unclear. Yes-associated protein (YAP) is one key mechanosensitive protein that controls MSC differentiation. Previous studies have related nuclear mechanics with YAP activity, but we still lack an understanding of what nuclear deformation specifically regulates YAP and its relationship with mechanical stimuli. Here, we report that maximum nuclear curvature is the most precise biophysical determinant for YAP mechanotransduction-mediated MSC differentiation and is a relevant parameter for stem cell-based therapies. We employed traction force microscopy and confocal microscopy to characterize the causal relationships between contractility and nuclear deformation in regulating YAP activity in MSCs. We observed that an increase in contractility compresses nuclei anisotropically, whereby the degree of asymmetric compression increased the bending curvature of the nuclear membrane. We then examined membrane curvature and tension using thin micropatterned adhesive substrate lines and an FRET-based tension sensor, revealing the direct role of curvature in YAP activity driven by both active and passive nuclear import. Finally, we employed micropatterned lines to control nuclear curvature and precisely direct MSC differentiation. This work illustrates that nuclear curvature subsumes other biophysical aspects to control YAP-mediated differentiation in MSCs and may provide a deterministic solution to some of the challenges in mesenchymal stem cell therapies.

Manipulating Stem Cell Fate with Disordered Bioactive Cues on Surfaces: The Role of Bioactive Ligand Selection.

The development of 2D or 3D bioactive platforms for rapidly isolating pure populations of cells from adult stem cells holds promise for advancing the understanding of cellular mechanisms, drug testing, and tissue engineering. Over the years, methods have emerged to synthesize bioactive micro- and nanostructured 2D materials capable of directing stem cell fate. We introduce a novel method for randomly micro- or nanopatterning any protein/peptide onto both 2D and 3D scaffolds via spray technology. Our goal is to investigate the impact of arranging bioactive micropatterns (ordered vs disordered) on surfaces to guide human mesenchymal stem cell (hMSC) differentiation. The spray technology efficiently coats materials with controlled, cost-effective bioactive micropatterns in various sizes and shapes. BMP-2 mimetic peptides were covalently grafted, individually or in combination with RGD peptides, onto activated polyethylene terephthalate (PET) surfaces through a spraying process, incorporating nano/microscale parameters like size, shape, and composition. The study explores different peptide distributions on surfaces and various peptide combinations. Four surfaces were homogeneously functionalized with these peptides (M1 to M4 with various densities of peptides), and six surfaces with disordered micro- and nanopatterns of peptides (S0 to S5 with different sizes of peptide patterns) were synthesized. Fluorescence microscopy assessed peptide distribution, followed by hMSC culture for 2 weeks, and evaluated osteogenic differentiation via immunocytochemistry and RT-qPCR for osteoblast and osteocyte markers. Cells on uniformly peptide-functionalized surfaces exhibited cuboidal forms, while those on surfaces with disordered patterns tended toward columnar or cuboidal shapes. Surfaces S4 and S5 showed dendrite-like formations resembling an osteocyte morphology. S5 showed significant overexpression of osteoblast (OPN) and osteocyte markers (E11, DMP1, and SOST) compared to control surfaces and other micropatterned surfaces. Notably, despite sharing an equivalent quantity of peptides with a homogeneous functionalized surface, S5 displayed a distinct distribution of peptides, resulting in enhanced osteogenic differentiation of hMSCs.

DRP1 inhibition-mediated mitochondrial elongation abolishes cancer stemness, enhances glutaminolysis, and drives ferroptosis in oral squamous cell carcinoma.

Mitochondrial dynamics play a fundamental role in determining stem cell fate. However, the underlying mechanisms of mitochondrial dynamics in the stemness acquisition of cancer cells are incompletely understood. Metabolomic profiling of cells were analyzed by MS/MS. The genomic distribution of H3K27me3 was measured by CUT&Tag. Oral squamous cell carcinoma (OSCC) cells depended on glucose or glutamine fueling TCA cycle were monitored by 13C-isotope tracing. Organoids and tumors from patients and mice were treated with DRP1 inhibitors mdivi-1, ferroptosis inducer erastin, or combination with mdivi-1 and erastin to evaluate treatment effects. Mitochondria of OSCC stem cells own fragment mitochondrial network and DRP1 is required for maintenance of their globular morphology. Imbalanced mitochondrial dynamics induced by DRP1 knockdown suppressed stemness of OSCC cells. Elongated mitochondria increased α-ketoglutarate levels and enhanced glutaminolysis to fuel the TCA cycle by increasing glutamine transporter ASCT2 expression. α-KG promoted the demethylation of histone H3K27me3, resulting in downregulation of SNAI2 associated with stemness and EMT. Significantly, suppressing DRP1 enhanced the anticancer effects of ferroptosis. Our study reveals a novel mechanism underlying mitochondrial dynamics mediated cancer stemness acquisition and highlights the therapeutic potential of mitochondria elongation to increase the susceptibility of cancer cells to ferroptosis.

PINK1 deficiency alters muscle stem cell fate decision and muscle regenerative capacity

Maintenance of mitochondrial function plays a crucial role in the regulation of muscle stem cell (MuSC), but the underlying mechanisms remain ill defined. In this study, we monitored mitophagy in MuSCS under various myogenic states and examined the role of PINK1 in maintaining regenerative capacity. Results indicate that quiescent MuSCs actively express mitophagy genes and exhibit a measurable mitophagy flux and prominent mitochondrial localization to autophagolysosomes, which become rapidly decreased during activation. Genetic disruption of Pink1 in mice reduces PARKIN recruitment to mitochondria and mitophagy in quiescent MuSCs, which is accompanied by premature activation/commitment at the expense of self-renewal and progressive loss of muscle regeneration, but unhindered proliferation and differentiation capacity. Results also show that impaired fate decisions in PINK1-deficient MuSCs can be restored by scavenging excess mitochondrial ROS. These data shed light on the regulation of mitophagy in MuSCs and position PINK1 as an important regulator of their mitochondrial properties and fate decisions.

Dynamic light-responsive RhoA activity regulates mechanosensitive stem cell fate decision in 3D matrices

The behavior of stem cells is regulated by mechanical cues in their niche that continuously vary due to extracellular matrix (ECM) remodeling, pulsated mechanical stress exerted by blood flow, and/or cell migration. However, it is still unclear how dynamics of mechanical cues influence stem cell lineage commitment, especially in a 3D microenvironment where mechanosensing differs from that in a 2D microenvironment. In the present study, we investigated how temporally varying mechanical signaling regulates expression of the early growth response 1 gene (Egr1), which we recently discovered to be a 3D matrix-specific mediator of mechanosensitive neural stem cell (NSC) lineage commitment. Specifically, we temporally controlled the activity of Ras homolog family member A (RhoA), which is known to have a central role in mechanotransduction, using our previously developed Arabidopsis thaliana cryptochrome-2-based optoactivation system. Interestingly, pulsed RhoA activation induced Egr1 upregulation in stiff 3D gels only, whereas static light stimulation induced an increase in Egr1 expression across a wide range of 3D gel stiffnesses. Actin assembly inhibition limited Egr1 upregulation upon RhoA activation, implying that RhoA signaling requires an actin-involved process to upregulate Egr1. Consistently, static-light RhoA activation rather than pulsed-light activation restricted neurogenesis in soft gels. Our findings indicate that the dynamics of RhoA activation influence Egr1-mediated stem cell fate within 3D matrices in a matrix stiffness-dependent manner.

Abstract 2580: Targeting chromatin in ovarian cancer stem cells

Abstract The dynamic organization of chromatin, governed by epigenetic modifiers, critically regulates cell plasticity and determines cell fate in both normal and malignant stem cells. We hypothesized that chromatin architecture plays a role in regulating transcriptional heterogeneity in ovarian cancer stem cells (OCSCs) to maintain stemness and chemoresistance. PWS microscopy showed that ALDH+ OCSCs derived displayed higher nuclear fractal dimension D signals (P<0.05) than ALDH- non-OCSCs, indicating that chromatin in OCSCs possesses higher packing compared to non-OCSCs. Cut & TAG Sequencing showed that of the 3606-histone repressive H3K27m3 enriched regions (P<0.05, fold change >2), 3208 were detected in OCSCs while 398 peaks were present in non-OCSCs, consistent with findings supporting high D features in OCSCs. The active mark H3K4me3 was enriched at the promoter of stemness-associated genes, including SOX2, ALDH1A3, and FZD7 in OCSCs; and the repressive histone mark H3K27me3 was enriched at the promoter of the epithelial cell marker CDH1 in OCSCs. These results support that the chromatin of OCSCs contains open genomic regions, corresponding to stemness-associated genes and repressed regions associated with genes related to differentiation. RNA sequencing indicated that the coefficient of variance (COV) reflecting transcriptional heterogeneity in OCSCs was higher than in non-OCSCs at baseline. In response to platinum, genes with low baseline expression in OCSCs were most upregulated, supporting increased transcriptional heterogeneity in response to chemotherapy. Inhibitors of epigenetic modifiers (DNMTi, EZH2i, and Dot1Li) induced chromatin re-organization in OCSCs and promoted OCSCs transition toward differentiation, supporting that stemness-associated chromatin structure is targetable. We demonstrated that a Dot1L inhibitor (Dot1Li) targeted OCSCs by decreasing chromatin packing D, resulting in decreased transcriptional heterogeneity. In conclusion, OCSCs harbor highly packed chromatin, contributing to chemoresistance and cell plasticity. Epigenetic modifiers (such as Dot1Li) inhibit stemness by regulating chromatin organization and transcriptional malleability. Citation Format: Yinu Wang, Jane Frederick, Yaqi Zhang, Elizabeth Thomas Bartom, Greta Bauer, Edward Tanner, Vadim Backman, Daniela Matei. Targeting chromatin in ovarian cancer stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2580.

ECM and epithelial stem cells: the scaffold of destiny.

Adult stem cells play a critical role in maintaining tissue homeostasis and promoting longevity. The intricate organization and presence of common markers among adult epithelial stem cells in the intestine, lung, and skin serve as hallmarks of these cells. The specific location pattern of these cells within their respective organs highlights the significance of the niche in which they reside. The extracellular matrix (ECM) not only provides physical support but also acts as a reservoir for various biochemical and biophysical signals. We will consider differences in proliferation, repair, and regenerative capacities of the three epithelia and review how environmental cues emerging from the niche regulate cell fate. These cues are transduced via mechanosignaling, regulating gene expression, and bring us to the concept of the fate scaffold. Understanding both the analogies and discrepancies in the mechanisms that govern stem cell fate in various organs can offer valuable insights for rejuvenation therapy and tissue engineering.

Deciphering "Immaturity-Stemness" in Human Epidermal Stem Cells at the Levels of Protein-Coding and Non-Coding Genomes: A Prospective Computational Approach.

The epidermis hosts populations of epithelial stem cells endowed with well-documented renewal and regenerative functions. This tissue thus constitutes a model for exploring the molecular characteristics of stem cells, which remain to date partially characterized at the molecular level in human skin. Our group has investigated the regulatory functions of the KLF4/TGFB1 and the MAD4/MAX/MYC signaling pathways in the control of the immaturity-stemness versus differentiation fate of keratinocyte stem and precursor cells from human interfollicular epidermis. We described that down-modulation of either KLF4 or MXD4/MAD4 using RNA interference tools promoted an augmented stemness cellular status; an effect which was associated with significant transcriptional changes, as assessed by RNA-sequencing. Here, we have implemented a computational approach aimed at integrating the level of the coding genome, comprising the transcripts encoding conventional proteins, and the non-coding genome, with a focus on long non-coding RNAs (lncRNAs). In addition, datasets of micro-RNAs (miRNAs) with validated functions were interrogated in view of identifying miRNAs that could make the link between protein-coding and non-coding transcripts. Putative regulons comprising both coding and long non-coding transcripts were built, which are expected to contain original pro-stemness candidate effectors available for functional validation approaches. In summary, interpretation of our basic functional data together with in silico biomodeling gave rise to a prospective picture of the complex constellation of transcripts regulating the keratinocyte stemness status.

Mechanical memory based on chromatin and metabolism remodeling promotes proliferation and smooth muscle differentiation in mesenchymal stem cells.

Stem cells respond and remember mechanical cues from the microenvironment, which modulates their therapeutic effects. Chromatin organization and energy metabolism regulate the stem cell fate induced by mechanical cues. However, the mechanism of mechanical memory is still unclear. This study aimed to investigate the effects of mechanical amplitude, frequency, duration, and stretch cycle on mechanical memory in mesenchymal stem cells. It showed that the amplitude was the dominant parameter to the persistence of cell alignment. F-actin, paxillin, and nuclear deformation are more prone to be remolded than cell alignment. Stretching induces transcriptional memory, resulting in greater transcription upon subsequent reloading. Cell metabolism displays mechanical memory with sustained mitochondrial fusion and increased ATP production. The mechanical memory of chromatin condensation is mediated by histone H3 lysine 27 trimethylation, leading to much higher smooth muscle differentiation efficiency. Interestingly, mechanical memory can be transmitted based on direct cell-cell interaction, and stretched cells can remodel the metabolic homeostasis of static cells. Our results provide insight into the underlying mechanism of mechanical memory and its potential benefits for stem cell therapy.